Kv2.1 channels, which are expressed in brain, heart, pancreas, and other organs and tissues, are important targets for drug design. Flecainide and propafenone are known to block Kv2.1 channels more potently than other Kv channels. Here, we sought to explore structural determinants of this selectivity. We demonstrated that flecainide reduced the K ؉ currents through Kv2.1 channels expressed in Xenopus laevis oocytes in a voltageand time-dependent manner. By systematically exchanging various segments of Kv2.1 with those from Kv1.2, we determined flecainide-sensing residues in the P-helix and inner helix S6. These residues are not exposed to the inner pore, a conventional binding region of open channel blockers. The flecainide-sensing residues also contribute to propafenone binding, suggesting overlapping receptors for the drugs. Indeed, propafenone and flecainide compete for binding in Kv2.1. We further used Monte Carlo-energy minimizations to map the receptors of the drugs. Flecainide docking in the Kv1.2-based homology model of Kv2.1 predicts the ligand ammonium group in the central cavity and the benzamide moiety in a niche between S6 and the P-helix. Propafenone also binds in the niche. Its carbonyl group accepts an H-bond from the P-helix, the amino group donates an H-bond to the P-loop turn, whereas the propyl group protrudes in the pore and blocks the access to the selectivity filter. Thus, besides the binding region in the central cavity, certain K ؉ channel ligands can expand in the subunit interface whose residues are less conserved between K ؉ channels and hence may be targets for design of highly desirable subtype-specific K ؉ channel drugs.Potassium channels are crucial in physiology. Medically important drugs block the open pore, which is formed by four subunits. Each subunit consists of transmembrane ␣-helical segments S1-S6 and the membrane re-entering extracellular P-loop. The pore-forming domain is composed of four S5-P-S6